CN109851605A - 葡萄籽中原花青素高聚体裂解与原花青素b3的提取方法 - Google Patents
葡萄籽中原花青素高聚体裂解与原花青素b3的提取方法 Download PDFInfo
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Abstract
本发明的公开的是葡萄籽中原花青素高聚体裂解与原花青素B3的提取方法，通过乙酸乙酯萃取，水相中原花青素高聚体共萃取三次，然后以儿茶素为裂解剂，乙醇为溶剂，在酸性下裂解反应，原花青素高聚体裂解为原花青素低聚体，本发明裂解反应条件可控，不但使高聚体得到利用，且增加了多酚的利用率，用色谱分离得到原花青素B3的含量，产物纯度高，达到99.1%，原花青素B3收率为3.5～4.8%。
Description
技术领域
本发明涉及葡萄籽中提取有效成份，具体涉及葡萄籽中原花青素高聚体裂解与原花青素B3的提取方法。
背景技术
葡萄籽提取物（Grape Seed Extract，GSE）是从葡萄籽中提取的一种人体内不能合成的新型高效天然抗氧化物质，它能有效清除人体内多余的自由基，具有延缓衰老和增强免疫力的作用。GSE中主要活性成分为葡萄籽多酚（Grape Proantho Cyanidins，GPC)，它由原花青素单聚体（儿茶素、表儿茶素、没食子酸酯）、低聚原花青素（Oligomeric ProanthoCyanidins，OPC)、高聚原花青素（Polymers Proantho Cyanidins，PPC)组成。三类原花青素聚合体体外抗氧化能力测试结果显示，随聚合度提高抗氧化能力增加。口服吸收实验研究结果显示，原花青素低聚体中二聚体（Procyanidins B1、B2、B3、B4）生物利用度最高，单聚体生物利用度较低，原花青素高聚体口服不吸收，葡萄籽原花青素高聚体需要裂解为低聚体才可以被生物吸收利用，因此若将原花青素高聚体裂解为原花青素低聚体中二聚体就提高了葡萄籽的利用率。
发明内容
本发明所要解决的技术问题是提供葡萄籽中原花青素高聚体裂解与原花青素B3的提取方法，该提取方法可以将原花青素高聚体裂解为原花青素低聚体，增加了原花青素B3提取率。
葡萄籽中原花青素高聚体裂解与原花青素B3的提取方法，其步骤如下：
a、将市售葡萄籽提取物10g溶于500ml水，4℃环境中静置分层，分离有机相，水相再用500ml乙酸乙酯萃取，萃取后分离，水相共萃取三次，最后水相减压浓缩至干燥，得黄色的高聚组分。有机相中为低聚组分，即多酚和原花青素低聚体，可以抗氧化物质利用；
b、将上述高聚组分溶于乙醇中，加儿茶素和滴加盐酸进行反应，盐酸终浓度为0.02～0.1M，反应温度控制在30～50℃，反应时间为10～30分钟，得反应液，所述高聚组分与所述乙醇的质量体积比为0.1g：5ml，所述高聚组分与所述儿茶素的质量比为0.1g：0.1～0.3g；
c、将上述反应液，用0.22μm滤膜过滤，液相色谱分离，色谱柱为Welch XB-C18，其规格为21.2mm×250mm、10μm，柱温25℃，流动相：乙腈和质量百分浓度0.2%的乙酸水，流动相中乙腈体积为20%，乙酸水体积为80%，等度洗脱，每次上样液50ml，检测波长280 nm，接收16～18min洗脱组分；
d、将上述洗脱组分，减压浓缩至干燥，与300～400目硅胶按重量比1:1混匀，过200～300目硅胶柱，乙酸乙酯洗脱，减压浓缩乙酸乙酯洗脱液至干燥，得到无色晶体原花青素B3。
本发明的优点在于葡萄籽中原花青素高聚体裂解与原花青素B3的提取方法，通过乙酸乙酯萃取，水相中原花青素高聚体共萃取三次，然后以儿茶素为裂解剂，乙醇为溶剂，在酸性下裂解反应，原花青素高聚体裂解为原花青素低聚体，本发明裂解反应条件可控，不但使高聚体得到利用，且增加了多酚的利用率，用色谱分离得到原花青素B3的含量，产物纯度高，达到99.1%，原花青素B3收率为3.5～4.8%。
具体实施方式
以下结合实施例对本发明作进一步详细描述。
实施例1
市售葡萄籽提取物10g溶于500ml水， 4℃环境中静置分层，水相用500ml乙酸乙酯萃取，萃取后分离，水相共萃取三次，水相萃取液浓缩至干燥，得黄色的高聚组分。有机相中为低聚组分，即多酚和原花青素低聚体，可以抗氧化物质利用；将高聚合度组分溶于乙醇中，加儿茶素和滴加盐酸，高聚组分与乙醇的质量体积比为0.1g：5ml，高聚组分与儿茶素的质量比为0.1g：0.2g，盐酸终浓度为0.04M，反应温度控制在35℃，反应时间22分钟，控制反应液过0.22μm滤膜过滤，滤液用色谱分离，色谱柱为Welch XB-C18，其规格为21.2mm×250mm、10μm，柱温25℃，流动相：乙腈与0.2%乙酸水的混合液，其中乙腈体积占20%，等度洗脱，检测波长280 nm，每次上样液50 ml，收集16～18 min洗脱组分，合并洗脱组分，减压浓缩至干燥，与300～400目硅胶按重量比1:1混匀，过200～300目硅胶柱，乙酸乙酯洗脱，减压浓缩乙酸乙酯相至干燥，得到0.279g无色晶体原花青素B3。
实施例2
与实施例1基本相同，所不同的只是高聚组分与儿茶素的质量比为0.1g：0.1g，盐酸终浓度为0.1M，反应温度控制在30℃，反应时间为30分钟。
实施例3
与实施例1基本相同，所不同的只是高聚组分与儿茶素的质量比为0.1g：0.3g，盐酸终浓度为0.02M，反应温度控制在50℃，反应时间为10分钟。
Claims (1)
1.葡萄籽中原花青素高聚体裂解与原花青素B3的提取方法，其特征在于步骤如下：
a、将市售葡萄籽提取物10g溶于500ml水，4℃环境中静置分层，分离有机相，水相再用500ml乙酸乙酯萃取，萃取后分离，水相共萃取三次，最后水相减压浓缩至干燥，得黄色的高聚组分；
b、将上述高聚组分溶于乙醇中，加儿茶素和滴加盐酸进行反应，盐酸终浓度为0.02～0.1M，反应温度控制在30～50℃，反应时间为10～30分钟，得反应液，所述高聚组分与所述乙醇的质量体积比为0.1g：5ml，所述高聚组分与所述儿茶素的质量比为0.1g：0.1～0.3g；
c、将上述反应液，用0.22μm滤膜过滤，液相色谱分离，色谱柱为Welch XB-C18，其规格为21.2mm×250mm、10μm，柱温25℃，流动相：乙腈和质量百分浓度0.2%的乙酸水，流动相中乙腈体积为20%，乙酸水体积为80%，等度洗脱，每次上样液50ml，检测波长280 nm，接收16～18min洗脱组分；
d、将上述洗脱组分，减压浓缩至干燥，与300～400目硅胶按重量比1:1混匀，过200～300目硅胶柱，乙酸乙酯洗脱，减压浓缩乙酸乙酯洗脱液至干燥，得到无色晶体原花青素B3。
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| CN101012216A (zh) * | 2007-02-02 | 2007-08-08 | 浙江大学 | 一种低聚体原花青素的制备方法 |
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| CN101012216A (zh) * | 2007-02-02 | 2007-08-08 | 浙江大学 | 一种低聚体原花青素的制备方法 |
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| Title |
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| NILS KOHLER等: "New Approach for the Synthesis and Isolation of Dimeric Procyanidins", 《J. AGRIC. FOOD CHEM》 * |
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